Toxic Effects of Formaldehyde and Solvents in Histology Technicians: A Preliminary Report

Abstract

Increased definition of cement lines aids in the identification of secondary osteons in bone. A staining protocol taken from Burr (Anat Rec 232:180–189, 1991) and modified by Havill using toluidine blue on plastic-embedded bone specimens was applied to cranial core and femoral shaft sections but proved to be too time-consuming and inconsistent. Experimentation resulted in a different protocol for each bone type, perhaps as the result of differences in section thickness and cortical area. Staining of femoral sections proved particularly difficult, with increased etching duration and agitation proving most influential in intensity and uniforlnity of stain. (The J Histotechnol 28:73, 2005)

Submitted August 20, 2004; accepted with revisions April 15, 2005     

甲苯胺蓝快速显示肥大细胞的制作方法探讨

目的为了使肥大细胞在标本制作过程中胞质内的颗粒不被水溶性液体所溶解,且使其标本上的颗粒颜色呈现异染性。方法选用20g以上的健康小白鼠为材料,用70%乙醇液为甲苯胺蓝染料的溶剂,一步完成标本的固定与染色操作。结果所显示的肥大细胞轮廓完整,与底衬背景对比鲜明,而且胞质内紫红色的嗜碱性颗粒粗大清晰、色彩鲜艳。结论在简化了操作程序的基础上,很好地保留了肥大细胞颗粒,又能使其颗粒染色呈现异染性。     

A Toluidine Blue Method for Demonstrating Mast Cells

Abstract

A modified Giemsa azure-eosin method for staining polymethyl methacrylate (PMMA) plastic embedded bone sections is described. The method is similar to those used to stain glycol methacrylate sections but differs in timing and addition of air drying steps. The stain yields effective differentiation of marrow cells, growth plate, calcified cartilage, osteoid, bone matrix and bone cells in undecalcified sections of bone. The stain does not interfere with the resolution of fluorochrome labeled specimens. Hence, the same slide can beused to evaluate static and dynamic parameters of bone metabolism. (The J Histotechnol 14:85, 1991)     

Observations on Fixation and Staining of Bone Marrow Biopsies

Abstract

Ten bone marrow (BM) biopsies were taken from the iliac crest of six autopsy cases. The tissues were fured in Zenker's fixative for intervals between one and 18 hours with and without decalcification. Ultrasonic bombardment was used for one and two hours. The same procedures were also used following 18 hours fixation in 10% neutral buffered formalin. Coded slides were double blind evaluated for section quality and cytologic detail after staining with H & E, three Giemsa variants, and Gomori's iron stain.

Paraffin processed BM biopsies were optimally prepared by fixation in Zenker's fluid for 4-18 hours without decalcification. Prior formalin fixation did not influence the consistent brilliance of Giemsa staining with the preferred Sheehan Wright Giemsa stain. Overnight Zenker fixation with or without prior formalin exposure and the absence of decalcification is recommended for the routine preparation of BM biopsies for paraffin processing. (The J of Histotechnol 11:145, 1988.)     

The Effect of Various Fixatives and Trypsin Digestion Upon the Staining of Routine Paraffin-Embedded Sections by the Peroxidase-Antiperoxidase and Immunofluorescent Technique

Abstract

The use of immunohistochemical techniques in biological and diagnostic pathology laboratories, and the theory of the unlabelled peroxidase-antiperioxidase technique, is briefly reviewed. The results of experiments using a variety of routine fixatives, with pretreatment of the sections with a trypsin solution, indicated that while Bouin's fixative alone was superior to formalin fixation, the pretreatment of sections with a trypsin solution gave markedly improved results with either fixative — the results with buffered formalin then being almost equal to those after Bouin's fixative. This methodology allows both prospective and retrospective immunohistochemical studies to be made on routinely fixed and processed paraplast-embedded tissues using the light microscope.     

A Method for Preparing Thin,Ground, Sections of Undecalcified Bone for Histomorphometry

Abstract

A method is described for obtaining thin, undecalcified, ground sections of bone. The procedure consisted of grinding gradually a plastic embedded 150uM bone section to 10uM thick using the Maruto Speed Lapping Machine ML-150 Dc. Sections prepared in this manner were suitable either for quantitative or qualitative bone histomorphometry.     

Alcian Blue Method for Demonstration of Histoplasma duboisii in Paraffin Sections

Abstract

A special staining technique for demonstrating Histoplasma duboisii in paraffin sections was developed during a study on the staining characteristics of this fungus, which acts as the causative organism for a type of fungal granuloma known as African histoplasmosis, found mostly in African countries. The method is relatively simple, reliable, and easily reproducible. It uses two common dyes, eosin and Alcian blue, that are readily available. The technique does not require any special fixatives and can be carried out on formalin fixed, parafin processed specimens. Because the method also eliminates the complicated and time-consuming special techniques previously used to demonstrate this fungus, it is more economical and time-saving. In paraffin sections, H. duboisii are demonstrated as round to ovoid sky-blue circlets in rose-pink background. (J Histotechnol 12:220, 1989).     

Preparation and Methods for Sequential Evaluation of Titanium Implant Interfaces in Plastic Embedded Calcified Bone

Abstract

Clinical, radiographic, fluorescence, histological, and electron microscopy techniques can be utilized for a comprehensive examination of bone/implant interfaces. To accomplish this objective, proper fixation, processing techniques, instrument quality control, and laboratory safety practices should be followed at all times. This paper reviews the methodology that has become standard practice in our laboratory for the evaluation of bone/irnplant interfaces. A regimen of fluorescent bone markers was used after implant placement to study rates of bone remodeling. At sacrifice, the tissue was perfused or immersed in an aldehyde or alcohol fixative. Implants were retrieved en bloc, radiographs made, and tissues embedded in plastic. Tissue implant blocks were cut longitudinally into 300–400 μm thick wafers with a low speed saw equipped with a diamond blade. The wafers were glued to plexiglass slides and hand ground to approximately 25 μm. Wafers were photographed with an epi-fluorescence microscope; radiographs were made in an X-ray cabinet; and the sections were stained for light microscopy evaluation. Various histomorphometric methods can be used to determine some variables of tissue remodeling around the implants from photomicrographs. Specimens prepared in this fashion can also be evaluated with a scanning or transmission electron microscope. These combined techniques provide a comprehensive approach for the evaluation of the bone/implant interface. (The J Histotechnol 17:333, 1994)     

Is it necessary to embed bone marrow biopsies in plastic for haematological diagnosis?

With the increase in the use of bone marrow trephines for diagnosis have come numerous reports that traditional methods of preparation (by decalcification and embedding in paraffin wax) should be replaced by plastic embedding to avoid decalcification. It has been argued that only by this means can the high quality preparations needed for accurate haematopathological diagnosis be achieved. The present study challenges this viewpoint and argues that with a little extra care and attention conventional paraffin embedding techniques can give equally high quality preparations. Sections prepared in this way meet the diagnostic needs of the haematologist, without requiring a separate technique to be established in the pathology laboratory solely for bone marrow trephines.     

Variation of the Holmes Method for Histologic Staining of Bone Canaliculi

Abstract

The Holmes method for soft tissue staining of nerve fibers was modified to produce excellent detail of the canalicular network in decalcified bone tissue. With simple changes in timing and the meticulous use of fresh reagents, we were able to stain bone lacunae and canaliculi in a variety of animal and human tissues. This technique produced more uniform and accurate results than the thionin/picric acid stains used previously for visualization of osteocytes. The morphologic results can be used for diagnostic purposes as well as stereological studies where the osteocyte network is of importance. (The J Histotechnol 16:355, 1993)     

Polymerization of Methacrylates Used as Histological Embedding Media: Local Variations and Time Course of Hardness of Methylmethacrylate Blocks

Abstract

Hardness of the blocks of polymerized methylmethacrylate (MMA, a plastic embedding medium widely used in calcified tissue histology) was measured at the top, the bottom, and the sides of the blocks. In addition, the most superficial layers of resins were removed with a grinding machine to explore the hardness in the inner parts of the blocks. A kinetic study of hardness changes was also done in all these areas during a period of 60 days. We observed an altered polymerization at the top of the blocks that we attribute to incompletely excluded atmospheric oxygen. However, a defect in the polymerization process was also observed on the lateral sides and the bottom of the blocks, suggesting complex changes induced by surface contact: MMA continued to harden quickly after blocks were been removed from the molds, and polymerization was not completed even if the blocks looked hard and suitable for sectioning. Heightening the tissue specimen with polyester foam or sponge (to avoid contact with the bottom of the mold) and waiting one or two days after removal from the mold were easy ways to avoid embedding, sectioning and staining artifacts.(The J Histotechnol 16:65, 1993)     

Osteo-radio-necrosis (ORN) and bisphosphonate-related osteonecrosis of the jaws (BRONJ): the histopathological differences under the clinical similarities

Objectives: Both Osteoradionecrosis (ORN) and Bisphosphonate associated osteonecrosis of the jaws (BRONJ) present clinically as regions of exposed necrotic bone. The study aimed to demonstrate the histopathological differences behind the observed clinical similarities. Study Design: Ten ORN specimens and ten BRONJ specimens were used, as well as ten samples of normal mandibular bone as control. Two bone-specific stainings were used, i.e. Sirius Red for the study of the relative presence of collagen types I and III and toluidine blue for the study the osteon density. Results: The Red Green Blue (RGB)-analysis of the specimens stained with Sirius Red identified significant differences between the chromatic patterns observed in bone preparations of patients suffering from ORN when compared to both BRONJ and control samples. Moreover, the osteon density of the BRONJ samples was significantly lower when compared to ORN and normal bone samples. Conclusions: The demonstrated differences in the bone architecture and in the bone collagen content between the two pathological conditions most likely reflect underlying pathophysiological differences.     

Immunohistochemical Method for the Simultaneous Demonstration of Three Proteins in EDTA Decalcified Paraffin Embedded Bone Sections

Abstract

Immunohistochemistry was used to demonstrate the presence of three proteins (actin, osteocalcin and alkaline phosphatase) within the same ethylenediaminetetraacetic acid (EDTA) decalcified and paraffin embedded bone tissue section. EDTA chelated calcium ions, causing endogenous alkaline phosphatase (ALP) to be inactivated. By ablating endogenous ALP activity through chelation, we were able to use an ALP based detection system before reactivation of the endogenous enzyme with magnesium chloride. After reactivation, endogenous ALP itself was demonstrated immunohistochemically. (The J Histotechnol 15:93, 1992)     

Microanatomical Record of Cortical Bone Remodeling and High Vascularity in a Fossil Giant Rat Midshaft Femur

Rat cortical bone does not typically undergo secondary (Haversian) remodeling. Haversian organization of rat bone has been mainly observed in experimental settings following biomechanical or dietary manipulation. Here, we report an observation of cortical secondary osteons within a histological femur cross-section from an extinct (late Quaternary) form of Timorese giant rat (Murinae gen. et sp. indet). The medio-lateral midshaft diameter of its femur, used as a measure of bone size, is 6.15 mm and indicates a heavier than normal skeletal frame. We compare this sample to bone histology in a small rat's midshaft femur of 2.33 mm diameter. A complete lack of Haversian bone remodeling characteristics is noted for the smaller sample, which is dominated by radial vascular canals. The giant rat shows clear secondary osteons and diffuse vascularity mainly composed of tightly packed longitudinal canals across its cortex. It appears that rat cortical bone can undergo bone remodeling, and is organized in a highly vascularized manner, in insular giant cases. Our findings from Timor align with results reported in experimental rat model skeletal biology literature and other insular fossil rat material. Where macroanatomical examination is limited, histological observations on fossil rat limb bones have the potential to aid reconstructions of life history and skeletal growth aspects in these rodents. Anat Rec, 302:1934–1940, 2019. © 2019 American Association for Anatomy     

豌豆骨移位代月骨的应用解剖

在15只成年上肢标本和57套腕骨标本上观察了豌豆骨的形态和血供。豌豆骨的骨内、外血管网发达,吻合丰富,有3～5支营养血管。以尺侧腕屈肌腱为蒂进行移位代替月骨,血供有保证。豌豆骨的纵径刚好与月骨的横径相近,故将纵置的豌豆骨改为横置方位:适当保留豌豆骨周围部分软组织,可以填补月骨空间,又能保存骨周围血管网。     

The Value of Histological Sections of Bone Marrow: Procedure and Special Stains for Evaluating Bone Marrow Sections

Abstract

Bone marrow biopsies, aspirates and smears are important procedures for diagnoses. Morphological abnormalities like focal lesions (lymphomas, granulomas) cannot be found in smears but will be present in the biopsy. Superior histologic preparations are essential for diagnostic interpretation of bone marrow. Appropriate histologic procedures for the preparation of bone marrow sections are presented in detail. Methods of fixation, processing, sectioning, routine H & E, iron stains and certain “special stains” are described.     

A Polychrome Stain for Mucopolysaccharides on Semithin Sections of Epoxy Resin Embedded Tissues

Abstract

The method describes a simple, quick polychromatic stain for Epon-Araldite and Araldite embedded tissue. It differentially stains mucins and mucopolysaccharides. The technique is simple to use and requires a hot plate to carry out the stain, which is done with droplets of Milipore filtered stain solution. It is useful as a screening stain or as a stain for some tissue components that generally require complex and prolonged procedures. A wide range of shades are produced, with the best effects achieved in tissues with abundant connective tissue, mucopolysaccharides, or granulocytic cells. Intraluminal bacteria of gastric biopsies are also demonstrated.     

Bone Microstructure of Pareiasaurs (Parareptilia) from the Karoo Basin,South Africa: Implications for Growth Strategies and Lifestyle Habits

Numerous morphological studies have been carried out on pareiasaurs; yet their taxonomy and biology remain incompletely understood. Earlier works have suggested that these herbivorous parareptiles had a short juvenile period as compared to the duration of adulthood. Several studies further suggested an (semi‐) aquatic lifestyle for these animals, but more recent investigations have proposed a rather terrestrial habitat. Bone paleohistology is regarded as a powerful tool to assess aspects of tetrapod paleobiology, but few studies have been conducted on pareiasaurs. The present study assesses intra and inter‐specific histovariability of pareiasaurs and provides fresh insights into their paleobiology, thereby permitting a re‐evaluation of earlier hypotheses. Our sample comprises various skeletal elements and several specimens covering most of the taxonomic and stratigraphic spectrum of South African pareiasaurs, including large and basal forms from the Middle Permian, as well as smaller and more derived forms from the Late Permian. Our results concerning size of elements and histological tissues show that for pareiasaurs, element size is not a good indicator of ontogenetic age, and furthermore, suggest that the specific diversity of the Middle Permian pareiasaurs may have been underestimated. The bone histology of these animals shows that they experienced a relatively rapid growth early in ontogeny. Periosteal growth later slowed down, but seems to have been protracted for several years during adulthood. Pareiasaur bone microanatomy is unusual for continental tetrapods, in having spongious stylopod diaphyses and thin compact cortices. Rigorous paleoecological interpretations are thus limited since no modern analogue exists for these animals. Anat Rec, 300:1039–1066, 2017. © 2016 Wiley Periodicals, Inc.     

Confocal laser scanning microscopy of porcine skin: implications for human wound healing studies

The structure of porcine skin as examined by light microscopy is reviewed and its similarities to and differences from human skin are highlighted. Special imaging techniques and staining procedures are described and their use in gathering morphological information in porcine skin is discussed. Confocal laser scanning microscopy (CLSM) was employed to examine the structure of porcine skin and the findings are presented as an adjunct to the information already available in the literature. It is concluded that CLSM provides valuable additional morphological information to material examined by conventional microscopy and is useful for wound healing studies in the porcine model.